Worldwide, millions of infants die of diarrhea each year, to say nothing of acute diarrheas in adults. All this is caused by E. coli of just a few of the many O-types. While rehydration therapy is generally the best temporary cure, prevention by vaccine administration is preferred for longer-term protection. This proposal touches upon a basic aspect of optimizing vaccine production that would not only be needed in fighting these diarrheal diseases but also useful generally in enterobacterial antigen productions for serological typing. In this line, recently this investigator, in collaboration with the International Escherichia Center (W.H.O.), has shown that culture temperature has marked effect on the size classes of lipopolysaccharide (LPS, O-antigen or endotoxin) of most of the O-types tested. Specifically, cells grown below 20 degrees C exhibit markedly higher average molecular weight LPS than cells grown at 37 degrees C. In addition, fewer low molecular weight components at the lower temperature are seen. AS whole cells or fractions thereof are commonly used as the source of antigen in immunization it is important to know what implications utilizing strains grown under various temperature regimes. It is proposed that cells of a number of selected 0-types that exhibit this thermal effect will be subjected firstly to a series of functional tests: relative sensitivities to phagocytosis, to serum killing, and to in vivo circulatory clearance and LD 50. Lastly, separately, both high and low temperature LPS's will be treated in a double cross experiment with the antibodies elicited by these antibodies to determine if the antigens differ in their epitopes depending on polymer length. The immunogenicity experiments will be conducted by the investigator at the World Health Organization's International Escherichia Center in Copenhagen, while the remainder will be conducted in the investigator's laboratory.